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PubMed | University of Pennsylvania, Autonomous University of Barcelona, University of Verona, University of Perugia and 9 more.
Type: Clinical Trial | Journal: British journal of haematology | Year: 2015

This study evaluated 65 pregnancies in 34 women with five different inherited platelet function disorders. Gestation was similar to that of the general population. Severe bleeds requiring blood transfusions were observed in 50% of deliveries in Glanzmann thrombasthenia (GT), but not in the patients with delta storage pool disease, Hermansky-Pudlak syndrome, P2Y12 defect or defect of thromboxane A2 receptor. Of note, severe haemorrhage also occurred in women with GT who had received prophylactic platelet transfusions, suggesting that better preventive treatments are required. Diagnosis and degree of spontaneous bleeding tendency before pregnancy were reliable parameters to predict the delivery-related bleeding risk.


Laguerre M.,CNRS Institute of Chemistry | Sabi E.,University of Sheffield | Daly M.,University of Sheffield | Stockley J.,University of Sheffield | And 4 more authors.
PLoS ONE | Year: 2013

Mutations in ITGA2B and ITGB3 cause Glanzmann thrombasthenia, an inherited bleeding disorder in which platelets fail to aggregate when stimulated. Whereas an absence of expression or qualitative defects of αIIbβ3 mainly affect platelets and megakaryocytes, αvβ3 has a widespread tissue distribution. Little is known of how amino acid substitutions of β3 comparatively affect the expression and structure of both integrins. We now report computer modelling including molecular dynamics simulations of extracellular head domains of αIIbβ3 and αvβ3 to determine the role of a novel β3 Pro189Ser (P163S in the mature protein) substitution that abrogates αIIbβ3 expression in platelets while allowing synthesis of αvβ3. Transfection of wild-type and mutated integrins in CHO cells confirmed that only αvβ3 surface expression was maintained. Modeling initially confirmed that replacement of αIIb by αv in the dimer results in a significant decrease in surface contacts at the subunit interface. For αIIbβ3, the presence of β3S163 specifically displaces an α-helix starting at position 259 and interacting with β3R261 while there is a moderate 11% increase in intra-subunit H-bonds and a very weak decrease in the global H-bond network. In contrast, for αvβ3, S163 has different effects with β3R261 coming deeper into the propeller with a 43% increase in intra-subunit H-bonds but with little effect on the global H-bond network. Compared to the WT integrins, the P163S mutation induces a small increase in the inter-subunit fluctuations for αIIbβ3 but a more rigid structure for αvβ3. Overall, this mutation stabilizes αvβ3 despite preventing αIIbβ3 expression. © 2013 Laguerre et al.


PubMed | University of Verona, Service de Nephrologie, IRCCS Policlinico San Matteo Foundation, Service de Biologie Clinique Secteur Hematologie and 36 more.
Type: Journal Article | Journal: Haematologica | Year: 2014

Pregnancy in women with inherited thrombocytopenias is a major matter of concern as both the mothers and the newborns are potentially at risk of bleeding. However, medical management of this condition cannot be based on evidence because of the lack of consistent information in the literature. To advance knowledge on this matter, we performed a multicentric, retrospective study evaluating 339 pregnancies in 181 women with 13 different forms of inherited thrombocytopenia. Neither the degree of thrombocytopenia nor the severity of bleeding tendency worsened during pregnancy and the course of pregnancy did not differ from that of healthy subjects in terms of miscarriages, fetal bleeding and pre-term births. The degree of thrombocytopenia in the babies was similar to that in the mother. Only 7 of 156 affected newborns had delivery-related bleeding, but 2 of them died of cerebral hemorrhage. The frequency of delivery-related maternal bleeding ranged from 6.8% to 14.2% depending on the definition of abnormal blood loss, suggesting that the risk of abnormal blood loss was increased with respect to the general population. However, no mother died or had to undergo hysterectomy to arrest bleeding. The search for parameters predicting delivery-related bleeding in the mother suggested that hemorrhages requiring blood transfusion were more frequent in women with history of severe bleedings before pregnancy and with platelet count at delivery below 50 10(9)/L.


Du L.M.,Medical College of Wisconsin | Du L.M.,Childrens Hospital of Wisconsin | Du L.M.,Fund Research Center | Nurden P.,Plateforme Technologique et dInnovation Biomedicale | And 29 more authors.
Nature Communications | Year: 2013

It is essential to improve therapies for controlling excessive bleeding in patients with haemorrhagic disorders. As activated blood platelets mediate the primary response to vascular injury, we hypothesize that storage of coagulation Factor VIII within platelets may provide a locally inducible treatment to maintain haemostasis for haemophilia A. Here we show that haematopoietic stem cell gene therapy can prevent the occurrence of severe bleeding episodes in dogs with haemophilia A for at least 2.5 years after transplantation. We employ a clinically relevant strategy based on a lentiviral vector encoding the ITGA2B gene promoter, which drives platelet-specific expression of human FVIII permitting storage and release of FVIII from activated platelets. One animal receives a hybrid molecule of FVIII fused to the von Willebrand Factor propeptide-D2 domain that traffics FVIII more effectively into a-granules. The absence of inhibitory antibodies to platelet-derived FVIII indicates that this approach may have benefit in patients who reject FVIII replacement therapies. Thus, platelet FVIII may provide effective long-term control of bleeding in patients with haemophilia A. © 2013 Macmillan Publishers Limited. All rights reserved.


Fang J.,Medical College of Wisconsin | Fang J.,Childrens Hospital of Wisconsin | Fang J.,Fund Research Center | Nurden P.,Plateforme Technologique et dInnovation Biomedicale | And 11 more authors.
Journal of Thrombosis and Haemostasis | Year: 2013

Background and objectives: β3-Deficient megakaryocytes were modified by human β3-lentivirus transduction and transplantation to express sufficient levels of a C560Rβ3 amino acid substitution, for investigation of how an activated αIIbβ3 conformation affects platelets in vivo in mice. Patient/Methods: As in our previous report of an R560β3 mutation in a patient with Glanzmann thrombasthenia, R560β3 murine platelets spontaneously bound antibody that only recognizes activated αIIbβ3 bound to its ligand, fibrinogen. Results: With this murine model, we showed that αIIb-R560β3 mutation-mediated continuous binding of fibrinogen occurred in the absence of P-selectin surface expression, indicating that the integrin was in an active conformation, although the platelets circulated in a quiescent manner. Remarkably, only 35% of R560β3 'mutant' mice survived for 6 months after transplantation, whereas 87% of C560β3 'wild-type' mice remained alive. Pathologic examination revealed that R560β3 mice had enlarged spleens with extramedullary hematopoiesis and increased hemosiderin, indicating hemorrhage. R560β3 megakaryocytes and platelets showed abnormal morphology and irregular granule distribution. Interestingly, R560β3 washed platelets could aggregate upon simultaneous addition of fibrinogen and physiologic agonists, but aggregation failed when platelets were exposed to fibrinogen before activation in vitro and in vivo. Conclusions: The results demonstrate that continuous occupancy of αIIbβ3 with fibrinogen disrupts platelet structure and function, leading to hemorrhagic death consistent with Glanzmann thrombasthenia rather than a thrombotic state. © 2013 International Society on Thrombosis and Haemostasis.


Nurden A.T.,Plateforme Technologique et dInnovation Biomedicale | Pillois X.,Plateforme Technologique et dInnovation Biomedicale | Pillois X.,French Institute of Health and Medical Research | Wilcox D.A.,Blood Research Institute | Wilcox D.A.,Medical College of Wisconsin
Seminars in Thrombosis and Hemostasis | Year: 2013

Glanzmann thrombasthenia (GT) is the principal inherited disease of platelets and the most commonly encountered disorder of an integrin. GT is characterized by spontaneous mucocutaneous bleeding and an exaggerated response to trauma caused by platelets that fail to aggregate when stimulated by physiologic agonists. GT is caused by quantitative or qualitative deficiencies of αIIbβ3, an integrin coded by the ITGA2B and ITGB3 genes and which by binding fibrinogen and other adhesive proteins joins platelets together in the aggregate. Widespread genotyping has revealed that mutations spread across both genes, yet the reason for the extensive variation in both the severity and intensity of bleeding between affected individuals remains poorly understood. Furthermore, although genetic defects of ITGB3 affect other tissues with β3 present as αvβ 3 (the vitronectin receptor), the bleeding phenotype continues to dominate. Here, we look in detail at mutations that affect (i) the β-propeller region of the αIIb head domain and (ii) the membrane proximal disulfide-rich epidermal growth factor (EGF) domains of β3 and which often result in spontaneous integrin activation. We also examine deep vein thrombosis as an unexpected complication of GT and look at curative procedures for the diseases, including allogeneic stem cell transfer and the potential for gene therapy. © 2013 by Thieme MedicalPublishers, Inc.


Nurden A.T.,Plateforme Technologique et dInnovation Biomedicale | Pillois X.,Plateforme Technologique et dInnovation Biomedicale | Pillois X.,French Institute of Health and Medical Research | Nurden P.,Plateforme Technologique et dInnovation Biomedicale
Expert Review of Hematology | Year: 2012

Glanzmann thrombasthenia (GT) is characterized by mucocutaneous bleeding due to platelets that fail to aggregate in response to physiologic stimuli. GT, a rare inherited disease, is caused by quantitative or qualitative deficiencies of αIIbβ3, an integrin receptor for adhesive proteins. Coded by the ITGA2B and ITGB3 genes, αIIbβ3 mediates platelet-to-platelet attachment, aggregation and clot retraction. Despite widespread mutation analysis, the reason for the extensive variation in both the severity and intensity of bleeding among affected individuals remains poorly understood. Although genetic defects of ITGB3 affect other tissues where β3 is present as αvβ3 (the vitronectin receptor), the bleeding phenotype continues to dominate. The authors now examine the relationship between genotype and phenotype in classic and variant forms of GT, and reassess if the nature of the gene mutation influences bleeding and treatment aimed at restoring hemostasis. © 2012 Expert Reviews Ltd.


Topalov N.N.,Center for Theoretical Problems of Physicochemical Pharmacology | Yakimenko A.O.,HemaCore LLC | Canault M.,Aix - Marseille University | Canault M.,French Institute of Health and Medical Research | And 13 more authors.
Arteriosclerosis, Thrombosis, and Vascular Biology | Year: 2012

OBJECTIVE-: Phosphatidylserine (PS) externalization by platelets upon activation is a key event in hemostasis and thrombosis. It is currently believed that strong stimulation of platelets forms 2 subpopulations, only 1 of which expresses PS. METHODS AND RESULTS-: Here, we demonstrate that physiological stimulation leads to the formation of not 1 but 2 types of PS-expressing activated platelets, with dramatically different properties. One subpopulation sustained increased calcium level after activation, whereas another returned to the basal low-calcium state. High-calcium PS-positive platelets had smaller size, high surface density of fibrin(ogen), no active integrin αIIbβ3, depolarized mitochondrial membranes, gradually lost cytoplasmic membrane integrity, and were poorly aggregated. In contrast, the low-calcium PS-positive platelets had normal size, retained mitochondrial membrane potential and cytoplasmic membrane integrity, and combined retention of fibrin(ogen) with active αIIbβ 3 and high proaggregatory function. Formation of low-calcium PS-positive platelets was promoted by platelet concentration increase or shaking and was decreased by integrin αIIbβ3 antagonists, platelet dilution, or in platelets from kindlin-3-deficient and Glanzmann thrombasthenia patients. CONCLUSION-: Identification of a novel PS-expressing platelet subpopulation with low calcium regulated by integrin αIIbβ3 can be important for understanding the mechanisms of PS exposure and thrombus formation. © 2012 American Heart Association, Inc.


PubMed | Plateforme Technologique et dInnovation Biomedicale
Type: | Journal: Case reports in hematology | Year: 2013

Patients with Glanzmann thrombasthenia fail to form large platelet thrombi due to mutations that affect the biosynthesis and/or function of the IIb3 integrin. The result is a moderate to severe bleeding syndrome. We now report unusual vascular behaviour in a 55-year-old woman with classic type I disease (with no platelet IIb3 expression) and a homozygous ITGA2B missense mutation (E324K) affecting the terminal -propeller domain of IIb. While exhibiting classic bleeding symptoms as a child, in later life this woman first developed deep vein thrombosis after a long air flight then showed vascular problems characteristic of Raynauds phenomenon, and finally this year she presented with chest pains suggestive of coronary heart disease. Yet while coronary angiography first showed a stenosis, this was not seen on a second examination when she was diagnosed with coronary spastic angina and Prinzmetal phenomenon. It is significant that the absence of platelet aggregation with physiologic agonists had not prevented any of the above cardiovascular or vascular diseases.


PubMed | Plateforme Technologique et dInnovation Biomedicale
Type: Journal Article | Journal: Seminars in thrombosis and hemostasis | Year: 2013

Glanzmann thrombasthenia (GT) is the principal inherited disease of platelets and the most commonly encountered disorder of an integrin. GT is characterized by spontaneous mucocutaneous bleeding and an exaggerated response to trauma caused by platelets that fail to aggregate when stimulated by physiologic agonists. GT is caused by quantitative or qualitative deficiencies of IIb3, an integrin coded by the ITGA2B and ITGB3 genes and which by binding fibrinogen and other adhesive proteins joins platelets together in the aggregate. Widespread genotyping has revealed that mutations spread across both genes, yet the reason for the extensive variation in both the severity and intensity of bleeding between affected individuals remains poorly understood. Furthermore, although genetic defects of ITGB3 affect other tissues with 3 present as v3 (the vitronectin receptor), the bleeding phenotype continues to dominate. Here, we look in detail at mutations that affect (i) the -propeller region of the IIb head domain and (ii) the membrane proximal disulfide-rich epidermal growth factor (EGF) domains of 3 and which often result in spontaneous integrin activation. We also examine deep vein thrombosis as an unexpected complication of GT and look at curative procedures for the diseases, including allogeneic stem cell transfer and the potential for gene therapy.

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